SUBMISSION GUIDELINES

PLEASE NOTE NEW VOLUME AND CONCENTRATION REQUIREMENTS

Samples can be submitted in PCR strip tubes or PCR plates for Sanger sequencing using the following process and guidelines.  A maximum of 92 samples can be submitted in a 96-well plate to allow for the inclusion of controls.  Your order request must be submitted online using dnaLIMS prior to physically submitting your samples:

Process

1

Prepare samples specific for the preferred service level, and follow the recommended protocol

SBC_Core-Prepared
User-Prepared

2

Enter your order request online using dnaLIMS, and submit samples with a printed copy of your request form together in a sealed bag.

 

3

Drop off in-person, by campus mail, or by courier to:

Sequencing and Bioinformatics Consortium
University of British Columbia
Room 3124 - PharmSci Building
2405 Wesbrook Mall
Vancouver, BC V6T 1Z3

Submission deadines: 9:30 am for SBC_Core-Prepared samples or 1:00 pm for all other submissions
Samples can be left in the refrigerated dropbox on the main floor of the Pharmaceutical Sciences building (contact us for details).   

 

Following these guidelines will ensure that your samples are processed without delay.

REMEMBER: Templates and primers should be resuspended in water or Tris (not TE), as EDTA will interfere with the ion concentration in the sequencing reaction. 

 

sanger__service_levels.jpg

 


 

Template and Primer Requirements

Template Considerations

Of primary importance to the success of DNA sequencing reactions is the quality and quantity of sample provided. SBC cannot be held responsible for failures in sequencing which are related to poor preparation of the DNA template used, or failure to follow recommended guidelines.

  • Contaminated template will give high background noise and poor, or no, sequence information
  • Many of the commercially-available extraction and purification kits (eg. Qiagen, Promega, Wizard Plus, Sigma) are recommended
  • *** Please ensure that the elution buffer does NOT contain EDTA ***
  • It is critical that the concentrations are measured carefully, as incorrect quantitation (whether higher or lower) will result in poor sequencing results. Please note that the required concentrations listed are based on UV absorbance / nanodrop and not fluorometric quantification. See also “How should I quantify my DNA prior to submission?” in our FAQ section: https://sequencing.ubc.ca/our-services-equipment/sanger-sequencing/sanger-faq
  • Modified conditions available for “difficult” templates (GC-rich, poly-T)

SBC_Core Prepared Requirements |  User-Prepared Requirements

Primer Considerations

Recommendations when designing custom sequencing primers:

  • Tm between 55°C and 60°C
  • Formula: ºC = 4(G+C) + 2(A+T)
  • “G” or “C” at 3’end
  • avoid primers with runs of 3 or more of the same base
  • use primer design software to avoid primers with hairpin or dimer characteristics

Custom primers must be submitted at a concentration of 5pmol/µl, resuspended in distilled water or 10 mM Tris pH 7-8, with no EDTA.

Standard Sequencing Primers available at SBC


Orientation Grid

 

96-well-plate.PNG

 

96-well-plate-orientation.png

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